Comparison of geneXpert versus sputum/gastric aspirate smear for AFB for the diagnosis of pulmonary tuberculosis in children

Introduction : This was a hospital based observational study for the comparison of geneXpert versus sputum /Gastric aspirate sample for AFB, for diagnosing pulmonary TB in children.The study was donein need for the useful diagnostic test, which is easy to perform, more sensitive and quicker. Gene Xpert assay is a single tube, cartridge based, real time PCR assay for the detection of tuberculosis. Materials and Methods: We enrolled 50 children for our study. Children attending inpatient or outpatient department were screened using a questionnaire. Suspected children were investigated with Complete Blood count, ESR, Chest Xray and Mantoux test. Sputum (in older children) was collected in a sterile container for AFB, and inspecialized container for geneXpert studies. In smaller children gastric aspirates were collected and sent for analysis and the results were compared. Results: Of the 50 children who were included in the study, 42 (84%) were less than 10 years, 8 (16%) were 11-16 years. 46% of patients had positive contact history of tuberculosis. History of clinical tuberculosis was present in 58% of the patients. Gene X pert analysis came positive in 42% of the patients whereas sputum/gastric aspirate analysis for AFB came positive in 12% of the patients. In our study, sensitivity, specificity, positive predictive value and negative predictive value of Gene Xpert analysis came as 28.6%, 100%, 100% and 65.9% with 70% accuracy. Conclusion: Gene Xpert analysis can be effectively used as a quick and accurate diagnostic test for the diagnosis of pulmonary tuberculosis in children, especially in a low resource setting, although the bacteriological culture remains the gold standard.


Introduction
Pediatric Tuberculosis has many diagnostic challenges such as-paucibacillary infection, absence of clear signs and symptoms, difficulty in obtaining good and adequate samples from children, difficulty in the interpretation of Mantoux test in younger children [1]. Most often, multiple tests are done to make a definitive diagnosis of TB. Some times children are treated empirically to prove TB infection retrospectively.
Hence there is a need for useful diagnostic test, which is easy toperform, more sensitive andquicker. Early diagnosis results in early initiation of treatment by RNTCP and hence better controlof tuberculosis in children.There are various diagnostic modalities for tuberculosis. Gene Xpert assay is a single tube, cartridge based, real time PCR assay for the detection of tuberculosis. The system allows simultaneous detection of both M.tuberculosis and rifampicin resistance. It is exceptionally sensitive for the detection of M.tuberculosis even in smear negative specimens. The result is available in 2 hours. Being cartridge based, the risk of cross contamination is less. It requires no in strumentation other than the GeneXpert system.
In 2010, WHO has endorsed the use of this assay as an initial test in all children with suspected TB, though bacteriological confirmation is the gold standard for the diagnosis. Gene Xpert assay is strongly recommended by WHO as an initial diagnostic test in children suspected of having MDR TB or HIV associated TB [2, 3,4]. This test detects both live and dead bacteria, hence not useful during follow up, except to detect rifampicin resistance. It can also be used to diagnose extrapulmonary tuberculosis depending on what specimen is sent for analysis.

Results
The present study was done to compare geneXpert analysis and sputum /gastric aspirate smear for AFB in order to diagnose pulmonary tuberculosis in children. Children presenting to paediatric unit/ outpatient department, satisfying the inclusion criteria were taken into the study. Total numbers of children included in the study were 50. Of these, 42 children (84%) were between 1-10 years, 8 children (16%) were 11-18 yrs as shown in Table 1.  Table 2 and Table 3.   This was statistically significant with a p value of 0.001. 6 (28.6%) children had positive sputum smear for AFB with significant p value of 0.003.
Therefore children with contact history and/or clinical history of tuberculosis are more likely to have tuberculosis. Sputum may or may not be positive in all of them, Sputum Positive 0(0%) 6(28.6%) 6(12%) 0.003** Table 7 explains sensitivity, specificity, PPV, NPV and accuracy of gene Xpert analysis. In our study sensitivity of Gene Xpert analysis was 28%, specificity was 100%, PPV was 100%, NPV was 65.9%, accuracy was 70%.

Discussion
The diagnosis and timely treatment of pulmonary tuberculosis in children is affected worldwide by the absence of quick and reliable tests. The gene Xpert analysis serves as useful rapid alternative test for the diagnosis of childhood TB [1].
Gene Xpert Assay is a single tube, cartridge based real time PCR assay for the detection of tuberculosis. It is exceptionally sensitive for the detection of mycobacterium TB even in smear negative specimen and the results are available in 2 hrs.
In our study of 50 patients, 42(84%) were between 1 -10 years of age, similar to the study done by Pang. Yet al [1] where percentage of children between 1-5 years of age enrolled for the study was significantly higher compared to the older age group. They concluded that younger children from low socioeconomic group have a higher risk for tuberculosis. Kumar et al observed similar results in his study [5]. 90.6% of the patients were< 10years of age. On the contrary, study done by Nhu et al [6], almost half of the children (47.9%) were between 11-16 years old. Only 31.5% of the children were 0-5 years of age.
Kumar A et al [5] found that contact history of tuberculosis was present only in 18.4% of cases and majority of the patients (81.6%) had no history of contact with tuberculosis patients. In our study, 46% of the patients had contact history of tuberculosis and 54% had no history of contact.
Our study also showed that contact history was positive in 23 children and 66% of them had Gene Xpert positive.
We can infer that patients end up contacting the disease, even without any history of contact with tuberculosis positive patient. It is also true that if contact history is positive, the chances of getting tuberculosis are higher.
In the study done by Rachow A et al, 164 children were studied. Gene Xpert analysis detected 100% of smear positive cases. Blood culture was also done and was compared with gene Xpert analysis. Gene Xpert analysis detected 3 fold more confirmed TB cases than smear microscopy. In a similar study done by Sekadde MP et al [9], 9% of smear positive, culture positive cases were identified by Gene Xpert analysis. The Gene X pert analysis identified twice as many cases as smear microscopy (79.4% Vs 41.2%). Our study showed similar results. Gene Xpert analysis detected more than 2 fold TB cases as compared to smear microcopy (71.4% Vs 28.65%) but sputum culture was not considered in our study.
Numbers of studies have demonstrated the utility of gene Xpert analysis in diagnosing pulmonary tuberculosis. In our study, the overall sensitivity, specificity, PPV, NPV of gene Xpert was 28.6%, 100%, 100% and 65.9% respectively. This is comparable with other studies.

Conclusion
Gene Xpert analysis can be effectively used as a quick and an accurate diagnostic test for the diagnosis of pulmonary tuberculosis in children, especially in a low resource setting.
Tuberculosis is more prevalent in younger age group. It has to be suspected and evaluated, in spite of contact history being negative.
In our study gene Xpert analysis detected more than 2 fold cases when compared to smear microscopy. This molecular diagnostic assay appears very promising in the diagnosis of childhood tuberculosis. Early diagnosis results in early initiation of treatment by RNTCP and hence better outcome of pulmonary tuberculosis in children.
What this study adds to the existing knowledge-This study reinforces the fact that gene Xpert analysis of sputum or gastric aspirate can be used as a quick and a reliable test in the diagnosis of pulmonary tuberculosis in children especially considering the difficult diagnostic modalities.
Limitation of the study-Our study had few limitations. Firstly sample size was small. Hence larger study with more number of children is required. Gene Xpert analysis was not compared with the gold standardblood/ sputum culture. Rifampicin resistance was not detected as it was included in the objective of the study.
Funding: Nil, Conflict of interest: None initiated, Perission from IRB: Yes